Clinical trials with immune modulatory agents have reduced the loss of C-peptide in patients with new onset Type 1 diabetes supporting the central role of immune mediated destruction of cells in the disease pathogenesis. However, the responses to these agents has been transient raising a question as to whether all of the destruction of cells is immune mediated or is also affected by environmental or genetic factors. This notion is supported by the observation that non-immunologic factors such as age and glucose control modify the responses to immune therapies and the kinetics of cell death. We hypothesize that effective treatment and prevention strategies need to address both the immune mediated and non-immune mediated factors that trigger cell death. However, our ability to address this issue is limited because we do not have a means of directly measuring cell death. The environmental factors, such as glucose, can affect cell function which has been the principle readout in clinical trials. In this proposal, we plan to use a novel assay of cell killing, involving detection of cell derived INS DNA to identify the rates of cell destruction in the progression of T1D and the effects of immune therapies on this process. We will test 3 interrelated hypotheses: First, that cell killing commences after immunologic initiation but prior to disease onset. Our preliminary data suggest that increased levels of cell killing can first be detected about 1 yr prior to clinical presentation. We will also determine whether epigenetic changes in the INS DNA can identify cell responses to stress during the progression of disease. Second, immune therapies have different effects on cell killing. Both recovery of dysfunctional cells and reduced killing may explain the recovery of cell function following metabolic stabilization but the different agents may not all have the same effects. Third, that environmental and genetic factors modify cell killing. We will use this measurement and the available metabolic, demographic, genetic, and clinical data from T1D prevention and intervention studies to identify the mechanisms of action of drugs and the factors that modify progression of the disease. Our studies represent a novel approach to identify factors that may modify responses to immune therapies and the progression of disease. They may develop new treatment strategies based on an improved understanding of the kinetics and mechanisms of disease pathogenesis.